Effect of <i>L</i>. <i>mali</i> APS1 on chromosomal aberration counts in 200 metaphase CHO-K1 cells.

Effect of L. mali APS1 on chromosomal aberration counts in 200 metaphase CHO-K1 cells.</p

Ames test results of <i>L</i>. <i>mali</i> APS1 powder using <i>Samonella typhimurium</i> strains TA97a, TA98, TA100, TA102, and TA1535.

Ames test results of L. mali APS1 powder using Samonella typhimurium strains TA97a, TA98, TA100, TA102, and TA1535.</p

AMOEBA Force Field Trajectories Improve Predictions of Accurate p<i>K</i>_a Values of the GFP Fluorophore: The Importance of Polarizability and Water Interactions

Precisely quantifying the magnitude, direction, and biological functions of electric fields in proteins has long been an outstanding challenge in the field. The most widely implemented experimental method to measure such electric fields at a particular residue in a protein has been through changes in pKa of titratable residues. While many computational strategies exist to predict these values, it has been difficult to do this accurately or connect predicted results to key structural or mechanistic features of the molecule. Here, we used experimentally determined pKa values of the fluorophore in superfolder green fluorescent protein (GFP) with amino acid mutations made at position Thr 203 to evaluate the pKa prediction ability of molecular dynamics (MD) simulations using a polarizable force field, AMOEBA. Structure ensembles from AMOEBA were used to calculate pKa values of the GFP fluorophore. The calculated pKa values were then compared to trajectories using a conventional fixed charge force field (Amber03 ff). We found that the position of water molecules included in the pKa calculation had opposite effects on the pKa values between the trajectories from AMOEBA and Amber03 force fields. In AMOEBA trajectories, the inclusion of water molecules within 35 Å of the fluorophore decreased the difference between the predicted and experimental values, resulting in calculated pKa values that were within an average of 0.8 pKa unit from the experimental results. On the other hand, in Amber03 trajectories, including water molecules that were more than 5 Å from the fluorophore increased the differences between the calculated and experimental pKa values. The inaccuracy of pKa predictions determined from Amber03 trajectories was caused by a significant stabilization of the deprotonated chromophore’s free energy compared to the result in AMOEBA. We rationalize the cutoffs for explicit water molecules when calculating pKa to better predict the electrostatic environment surrounding the fluorophore buried in GFP. We discuss how the results from this work will assist the prospective prediction of pKa values or other electrostatic effects in a wide variety of folded proteins

Effect of <i>L</i>. <i>mali</i> APS1 on developmental toxicity study in maternal rats showing an overview of the effects on the main observed reproductive toxicity.

Effect of L. mali APS1 on developmental toxicity study in maternal rats showing an overview of the effects on the main observed reproductive toxicity.</p

Positive control of Ames test.

Positive control of Ames test.</p

Changes of micronucleus counts in peripheral blood of ICR mice treated with <i>L</i>. <i>mali</i> APS1 powder.

Changes of micronucleus counts in peripheral blood of ICR mice treated with L. mali APS1 powder.</p

Incidence of fetal abnormalities after administration of <i>L</i>. <i>mali</i> APS1 to maternal rats (F₀).

Incidence of fetal abnormalities after administration of L. mali APS1 to maternal rats (F0).</p

Additional file 1 of Predicting frailty in older adults using vocal biomarkers: a cross-sectional study

Additional file 1: Supplementary Table S1. A one-way analysis of variance on acoustic features differences, 2020. Supplementary Table S2. Sex-specific differences in the association between acoustic features and the probability of frailty among older adults, by the three frailty indices, 2020

Trichostatin A Modulates Thiazolidinedione-Mediated Suppression of Tumor Necrosis Factor α-Induced Lipolysis in 3T3-L1 Adipocytes

<div><p>In obesity, high levels of tumor necrosis factor α (TNFα) stimulate lipolysis in adipocytes, leading to hyperlipidemia and insulin resistance. Thiazolidinediones (TZDs), the insulin-sensitizing drugs, antagonize TNFα-induced lipolysis in adipocytes, thereby increasing insulin sensitivity in diabetes patients. The cellular target of TZDs is peroxisome proliferator-activated receptor γ (PPARγ), a nuclear receptor that controls many adipocyte functions. As a transcription factor, PPARγ is closely modulated by coregulators, which include coactivators and corepressors. Previous studies have revealed that in macrophages, the insulin-sensitizing effect of PPARγ may involve suppression of proinflammatory gene expression by recruiting the corepressor complex that contains corepressors and histone deacetylases (HDACs). Therefore, we investigated whether the corepressor complex is involved in TZD-mediated suppression of TNFα-induced lipolysis in 3T3-L1 adipocytes. Trichostatin A (TSA), a pan HDAC inhibitor (HDACI) that inhibits class I and II HDACs, was used to examine the involvement of HDACs in the actions of TZDs. TSA alone increased basal lipolysis and attenuated TZD-mediated suppression of TNFα-induced lipolysis. Increased basal lipolysis may in part result from class I HDAC inhibition because selective class I HDACI treatment had similar results. However, attenuation of TZD-mediated TNFα antagonism may be specific to TSA and related hydroxamate-based HDACI rather than to HDAC inhibition. Consistently, corepressor depletion did not affect TZD-mediated suppression. Interestingly, TSA treatment greatly reduced PPARγ levels in differentiated adipocytes. Finally, extracellular signal-related kinase 1/2 (ERK1/2) mediated TNFα-induced lipolysis, and TZDs suppressed TNFα-induced ERK phosphorylation. We determined that TSA increased basal ERK phosphorylation, and attenuated TZD-mediated suppression of TNFα-induced ERK phosphorylation, consistent with TSA’s effects on lipolysis. These studies suggest that TSA, through down-regulating PPARγ, attenuates TZD-mediated suppression of TNFα-induced ERK phosphorylation and lipolysis in adipocytes.</p></div

TSA attenuates Rosi-mediated suppression of TNFα-induced ERK1/2 phosphorylation in 3T3-L1 adipocytes.

<p>(A and B) 3T3-L1 adipocytes were pretreated with vehicle (DMSO) or 660 nM TSA, together with or without 1 µM Rosi (Rosi) for 24h. Cells were then treated with or without 10 ng/ml TNFα for 30 min. Cellular proteins were solubilized and subjected to SDS-PAGE and Western analysis with the indicated antibodies. Representative immunoblots and quantification data from five independent experiments are shown in 5A and B, respectively. (C) ERK phosphorylation correlates highly with lipolysis in the treatments of Rosi, TNFα, or both in the presence or absence of TSA, as shown by fitting with linear regression. Individual values were obtained from the experiments described in Figures. 1 and 7B.</p